STRUCTURAL STUDIES ON AMINOGLYCOSIDE PHOSPHOTRANSFERASES

这个子项目是许多利用资源的研究子项目之一 由NIH/NCRR资助的中心拨款提供。子项目的主要支持 而子项目的主要调查员可能是由其他来源提供的， 包括其它NIH来源。 列出的子项目总成本可能 代表子项目使用的中心基础设施的估计数量， 而不是由NCRR赠款提供给子项目或子项目工作人员的直接资金。 抗生素耐药性出现的一个主要因素是存在化学修饰常见抗生素的细菌酶。氨基糖苷类抗生素（如卡那霉素、妥布霉素和庆大霉素）就是目前几乎普遍耐药的一类抗菌药物。肠球菌对庆大霉素的高水平耐药是由APH（2？）在抗生素上的特定羟基上磷酸化的酶的亚家族，使用ATP作为共底物。了解这些酶如何结合和失活的氨基糖苷类将提供有价值的信息，这些酶的特异性抑制剂的设计。 我们正在研究所有四个APH（2？）磷酸转移酶，APH（2？）- Ia和APH（2？）- IIa来自屎肠球菌，APH（2？）- IIIa来自E. gallinarum和APH（2？）IVa来自E.石蒜黄APH（2？）-庆大霉素二元复合物和AMPPCP-链霉素三元复合物的三维结构IIa的结晶条件，并确定了APH（2？）- IIIa和APH（2？）- IVa建立。APH（2？）- IIIa和APH（2？）- IVa结构已被解决。 研究将集中在解决这些酶的二元和三元复合物的结构。

This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. Primary support for the subproject and the subproject's principal investigator may have been provided by other sources, including other NIH sources. The Total Cost listed for the subproject likely represents the estimated amount of Center infrastructure utilized by the subproject, not direct funding provided by the NCRR grant to the subproject or subproject staff. A major factor in the emergence of antibiotic resistance is the existence of bacterial enzymes that chemically modify common antibiotics. One such family of anti-bacterials to which there is now almost universal resistance are the aminoglycosides (e.g. kanamycin, tobramycin and gentimicin). High level resistance to gentamicin in enterococci is mediated by a group of four phosphotransferases belonging to the APH(2?) sub-family of enzymes which phosphorylate at a specific hydroxyl group on the antibiotic, using ATP as a cosubstrate. An understanding of how these enzymes bind and deactivate the aminoglycosides will provide valuable information for the design of specific inhibitors of these enzymes. We are studying all four of the APH(2?) phosphotransferases, APH(2?)-Ia and APH(2?)-IIa from Enterococcus faecium, APH(2?)-IIIa from E. gallinarum, and APH(2?)-IVa from E. casseliflavus. The three dimensional structures of the binary gentamicin complex and a ternary AMPPCP-streptomycin complex of APH(2?)-IIa have been determined and published and the crystallization conditions for APH(2?)-IIIa and APH(2?)-IVa established. Both the APH(2?)-IIIa and APH(2?)-IVa structures have been solved. Studies will focus on solving structures of binary and ternary complexes of these enzymes.